The present invention relates to novel compounds which inhibit IMPDH, and to methods of making such compounds. The invention also encompasses pharmaceutical compositions comprising these compounds. The compounds and pharmaceutical compositions of the invention are particularly well suited for inhibiting IMPDH enzyme activity and, consequently, may be advantageously used as therapeutic agents for IMPDH-associated disorders. This invention also relates to methods for inhibiting the activity of IMPDH using the compounds of this invention and related compounds.
Inosine monophosphate dehydrogenase (IMPDH) has been shown to be a key enzyme in the regulation of cell proliferation and differentiation. Nucleotides are required for cells to divide and replicate. In mammals, nucleotides may be synthesized through one of two pathways: the de novo synthesis pathway or the salvage pathway. The extent of utilization of each pathway is dependent on the cell type. This selectivity has ramifications with regard to therapeutic utility as described below.
IMPDH is involved in the de novo synthesis of guanosine nucleotides. IMPDH catalyzes the irreversible NAD-dependent oxidation of inosine-5xe2x80x2-monophosphate (xe2x80x9cIMPxe2x80x9d) to xanthosine-5xe2x80x2-monophosphate (xe2x80x9cXMPxe2x80x9d), Jackson et al., Nature 256:331-333 (1975).
IMPDH is ubiquitous in eukaryotes, bacteria and protozoa. The prokaryotic forms share 30-40% sequence identity with the human enzyme.
Two distinct cDNA""s encoding IMPDH have been identified and isolated. These transcripts are labeled type I and type II and are of identical size (514 amino acids). Collart et al., J. Biol. Chem. 263:15769-15772 (1988); Natsumeda et al., J. Biol. Chem. 265:5292-5295 (1990); and U.S. Pat. No. 5,665,583 to Collart et al. These isoforms share 84% sequence identity. IMPDH type I and type II form tetramers in solution, the enzymatically active unit.
B and T-lymphocytes depend on the de novo, rather than salvage pathway, to generate sufficient levels of nucleotides necessary to initiate a proliferative response to mitogen or antigen. Due to the B and T cell""s unique reliance on the de novo pathway, IMPDH is an attractive target for selectively inhibiting the immune system without also inhibiting the proliferation of other cells.
Immunosuppression has been achieved by inhibiting a variety of enzymes. Examples include: phosphatase calcineurin (inhibited by cyclosporin and FK-506); dihydroorotate dehydrogenase (DHODase), an enzyme involved in the biosynthesis of pyrimidines (inhibited by leflunomide and brequinar); the kinase FRAP (inhibited by rapamycin); and the heat shock protein hsp70 (inhibited by deoxyspergualin).
Inhibitors of IMPDH have also been described in the art. WO 97/40028 and U.S. Pat. No. 5,807,876 describe a class of urea derivatives that possess a common urea backbone. A large number of compounds are described in WO 97/40028 and U.S. Pat. No. 5,807,876, but several of the compounds suffer from drawbacks such as inferior solubility. A recent publication, WO 98/40381, describes a series of heterocyclic substituted anilines as inhibitors of IMPDH.
U.S. Pat. Nos. 5,380,879 and 5,444,072 and PCT publications WO 94/01105 and WO 94/12184 describe mycophenolic acid (xe2x80x9cMPAxe2x80x9d) and some of its derivatives as potent, uncompetitive, reversible inhibitors of human IMPDH type I and type II. MPA has been demonstrated to block the response of B and T-cells to mitogen or antigen. Immunosuppressants, such as MPA and derivatives of MPA, are useful drugs in the treatment of transplant rejection and autoimmune disorders, psoriasis, inflammatory diseases, including, rheumatoid arthritis, tumors and for the treatment of allograft rejection. These are described in U.S. Pat. Nos. 4,686234, 4,725622, 4,727,069, 4,753,935, 4,786,637, 4,808,592, 4,861,776, 4,868,153, 4,948,793, 4,952,579, 4,959,387, 4,992,467; 5.247,083; and U.S. patent application Ser. No. 07/927,260, filed Aug. 7, 1992. MPA does display undesirable pharmacological properties, such as gastrointestinal toxicity and poor bioavailability.
Tiazofurin, ribavirin and mizoribine also inhibit IMPDH. These nucleoside analogs are competitive inhibitors of IMPDH, however these agents inhibit other NAD dependent enzymes. This low level of selectivity for IMPDH limits the therapeutic application of tiazofurin, ribavirin and mizoribine. Thus, new agents which have improved selectivity for IMPDH would represent a significant improvement over the nucleoside analogs.
Mycophenolate mofetil, sold under the trade name CELLCEPT, is a prodrug which liberates MPA in vivo. It is approved for use in preventing acute renal allograft rejection following kidney transplantation. The side effect profile limits the therapeutic potential of this drug. MPA is rapidly metabolized to the inactive glucuronide in vivo. In humans, the blood levels of glucuronide exceed that of MPA. The glucuronide undergoes enterohepatic recycling causing accumulation of MPA in the bile and subsequently in the gastrointestinal tract. This together with the production of the inactive glucuronide effectively lowers the drug""s in vivo potency, while increasing its undesirable gastrointestinal side effects.
Unlike type I, type II mRNA is preferentially upregulated in human leukemic cell lines K562 and HL-60. Weber, J. Biol. Chem. 266: 506-509 (1991). In addition, cells from human ovarian tumors and leukemic cells from patients with chronic granulocytic, lymphocytic and acute myeloid leukemias also display an up regulation type II mRNA. This disproportionate increase in IMPDH activity in malignant cells may be addressed through the use of an appropriate IMPDH inhibitor. IMPDH has also been shown to play a role in the proliferation of smooth muscle cells, indicating that inhibitors of IMPDH may be useful in preventing restenosis or other hyperproliferative vascular diseases.
IMPDH has been shown to play a role in viral replication in some viral cell lines. Carr, J. Biol. Chem. 268:27286-27290 (1993). The IMPDH inhibitor VX-497, is currently being evaluated for the treatment of hepatitis C virus in humans. Ribavirin has also been used in the treatment of hepatitis C and B viruses and when used in combination with interferon an enhancement in activity was observed. The IMPDH inhibitor ribavirin is limited by its lack of a sustained response in monotherapy and broad cellular toxicity.
There remains a need for potent selective inhibitors of IMPDH with improved pharmacological properties, physical properties and fewer side effects. Such inhibitors would have therapeutic potential as immunosuppressants, anti-cancer agents, anti-vascular hyperproliferative agents, antiinflammatory agents, antifungal agents, antipsoriatic and anti-viral agents. The compounds of the present invention differ from those taught by the prior art and are effective inhibitors of IMPDH.
The present invention provides heterocyclic compounds of the following formula (I), their enantiomers, diastereomers, tautomers and pharmaceutically acceptable salts, prodrugs and solvates thereof, for use as IMPDH inhibitors: 
wherein:
X is a monocyclic or bicyclic ring system optionally containing up to 4 heteroatoms selected from N, O, and S, and wherein a CH2 adjacent to any of the said N, O or S heteroatoms is optionally substituted with oxo (xe2x95x90O), and wherein X is optionally substituted by 0-5 substituents chosen from A, R1, or R2;
A is R3 or R4;
R3 is a 5- or 6-membered heterocyclic ring system containing up to 4 heteroatoms selected from N, O, and S, said heterocyclic ring system being optionally substituted with 0-3 R5, wherein when R5 is hydroxy, the heterocycle may undergo tautomerization to an oxo species, or exist as an equilibrium mixture of both tautomers;
R4 is selected from H, F, Cl, Br, I, NO2, CF3, C0-C4 alkylCN, C1-C4alkoxy-, C0-C4 alkylhydroxy, C1-C4 alkyl-, C1-C4 alkylcarbonyl-, C0-C4 alkylOCOR6, C0-C4 alkylOC(xe2x95x90O)OR6, C0-C4 alkylOC(xe2x95x90O)NR6R7, NH2, NHR6, C0-C4 alkylNR6R7, C0-C4 alkylNR7C(xe2x95x90O)OR6, C0-C4 alkylNR6SO2NR6R7, C0-C4 alkylNR7SO2R6, C0-C4 alkylSR6, C0-C4 alkylS(O)R6, C0-C4 alkylSO2R6, SO3R7, C0-C4 alkylSO2NR6R7, C0-C4alkyl SO2NR7 CO(CR9R10)qR6, C0-C4 alkylCO2H, C0-C4 alkylCO2R6, C0-C4 alkylCONR6R7, and C0-C4CONR7SO2(CR9R10)qR6;
R5 is selected from H, halogen, NO2, C1-C4alkyl, C3-C10cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, oxo, C1-C4alkoxy, C1-C4alkylcarbonyl, CN, NH2, NHR6, NR6R7, SR7, S(O)R7, SO2R7, SO3R7, SO2NR6, CO2H, CO2R6, and CONR6R7;
R is H or C1-C4alkyl;
R1 and R2 are each independently selected from H, halogen, NO2, C1-C4alkyl, C3-C10cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, C1-C4alkoxy-, OR6, O(CR9R10)rCO2R6, O(CR9R10)m NR6R7, O(CR9R10)pCN, O(CR9R10)rC(xe2x95x90O)NR6R7, C1-C4alkylcarbonyl-, CN, NH2, NHR6, NR6R7, NR7(CR9R10)rCO2R6, NR7OR6, NR7(CR9R10)mOR6, NR7CH[(CR9R10)pOR6]2, NR7C[(CR9R10)pOR6]3, NR7C(xe2x95x90O)R6, NR7(CR9R10)mOR6, NR7(CR9R10)mNR6R7, NR7(CR9R10)mSO2(CR9R10)qR, SR7, S(O)R7, SO2R7, SO2NR6, SO3R7, SO2NR6, CO2H, CO2R6, and CONR6R7; or, alternatively, R1 and R2, when on adjacent carbon atoms, may be taken together to be methylenedioxy or ethylenedioxy;
R6, R7 and R8 are each independently selected from H, C1-C6alkyl, C3-C10 cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, C1-C6 alkylcarbonyl, C3-C7 cycloalkyl(C1-C5 alkyl)carbonyl, C1-C6 alkoxycarbonyl, aryl(C0-C5 alkyl)carbonyl, aryl(C1-C5 alkyloxy)carbonyl, heterocyclic(C0-C5 alkyl)carbonyl, heterocyclic(C1-C5 alkoxy)carbonyl, C1-C6alkylsulfonyl, aryllsulfonyl, heteroarylsulfonyl, C0-C4alkylaryl, C0-C4alkylheterocyclic, wherein said cycloalkyl, aryl or heterocyclic groups are substituted with 0-2 substituents independently selected from the group consisting of C1-C4alkyl, hydroxy, C1-C4 alkoxy, F, Cl, Br, haloalkyl, NO2 and CN;
or, alternatively, R6 and R7, or R6 and R8, or R7 and R8, when both substituents are on the same nitrogen atom [as in (xe2x80x94NR6R7) or (xe2x80x94NR7R8)], can be taken together with the nitrogen atom to which they are attached to form a heterocycle selected from 1-aziridinyl, 1-azetidinyl, 1-piperidinyl, 1-morpholinyl, 1-pyrrolidinyl, thiamorpholinyl, thiazolidinyl, 1-piperazinyl, 1-imidazolyl, 3-azabicyclo[3,2,2]nonan-3-yl, and 1-tetrazolyl, the said heterocycle being optionally substituted with 0-3 groups selected from oxo, C0-C4alkylOH, C0-C4alkylOC1-C4alkyl, C0-C4alkylCONH2, C0-C4alkylCO2C0-C4alkyl, C1-C6 alkyl, C1-C4 alkoxy, C3-C7 cycloalkyl, xe2x80x94C0-C6 alkylcarbonyl, C3-C7 cycloalkylcarbonyl, C1-C6 alkoxycarbonyl, C3-C7 cycloalkoxycarbonyl, xe2x80x94NHCOalkyl, aryl, heteroaryl, aryl alkoxycarbonyl, heteroaryl alkoxycarbonyl, C1-C6 alkylsulfonyl, arylsulfonyl and heteroarylsulfonyl;
B is a monocyclic or bicyclic ring system optionally containing up to 4 heteroatoms selected from N, O, and S, and wherein a CH2 adjacent to any of the said N, O or S heteroatoms is optionally substituted with oxo (xe2x95x90O), and wherein B is optionally substituted by one to four R11 groups;
D is a monocyclic or bicyclic ring system optionally containing up to 4 heteroatoms selected from N, O, and S, and wherein a CH2 adjacent to any of the said N, O or S heteroatoms is optionally substituted with oxo (xe2x95x90O), and wherein D is optionally substituted by one to four (CR9R10)n E groups;
n is an integer having a value from 0-4;
m is an integer having a value from 2-6;
p is an integer having a value from 1-3;
q is an integer having a value from 0-3;
r is an integer having a value from 0-6;
R9 is H or C1-C4alkyl;
R10 is selected from H or C1-C4 alkyl, C1-C4 alkylhydroxy, C1-C4alkylaryl or C1-C4alkylheteroaryl, wherein said aryl or heteroaryl group may be substituted with 0-3 groups independently selected from H, halogen, NO2, C1-C4alkyl, C3-C10cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, C1-C4alkoxy, C1-C4alkylcarbonyl, CN, NH2, NR6R7, SR6, S(O)R6, SO2R6, SO3R6, SO2NR6, CO2H, CO2R6, and CONR6R7;
R11 is selected from H, halogen, NO2, C1-C4alkyl, C3-C10cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, C1-C4alkoxy-, OR6, O(CR9R10)rCO2R6, O(CR9R10)mNR6R7, O(CR9R10)pCN, O(CR9R10)rC(xe2x95x90O)NR6R7, C1-C4alkylcarbonyl-, CN, NH2, NHR6, NR6R7, NR7(CR9R10)rCO2R6, NR7OR6, NR7(CR9R10)mOR6, NR7CH[(CR9R10)pOR6]2, NR7C[(CR9R10)pOR6]3, NR7C(xe2x95x90O)R6, NR7(CR9R10)mOR6, NR7(CR9R10)mNR6R7, NR7(CR9R10)mSO2(CR9R10)qR6, SR7, S(O)R7, SO2R7, SO2NR6, SO3R7, CO2H, CO2R6, and CONR6R7;
E is selected from H, halogen, NO2, C1-C4alkyl, C3-C10cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, haloalkyl, haloalkoxy, OH, OR6, CN, CHO, CO2R6, CONR6R7, OCOR6, OC(xe2x95x90O)OR6, OC(xe2x95x90O)NR6R7, OCH2CO2R6, C(xe2x95x90O)R6, NH2, NHR6, NR6R7, NR7C(xe2x95x90O)R6, NR7C(xe2x95x90O)OR6, NR7C(xe2x95x90O)C(xe2x95x90O)OR6, NR7C(xe2x95x90O)C(xe2x95x90O)NR6R7, NR7C(xe2x95x90O)C(xe2x95x90O)(C1-C6alkyl), NR7C(xe2x95x90NCN)OR6, NR7C(xe2x95x90O)NR6R7, NR7C(xe2x95x90NCN)NR6R7, NR7C(xe2x95x90NR6)NR7R8, NR6SO2NR6R7, NR7SO2R6, SR6, S(xe2x95x90O)R6, SO2R6, SO3R7, SO2NR6R7, NHOH, NHOR6, NR6NR7NR8, N(COR6)OH, N(CO2R6)OH, CO2R6, CONR6R7, CONR7(CR9R10)rR6, CO(CR9R10)pO(CHR9)qCO2R6, CO(CR9CR10)rOR6, CO(CR9R10)pO(CR9R10)qR6, CO(CR9CR10)rNR6R7,OC(O)O(CR9R10)mNR6R7, O(CO)N(CR9R10)rR6, O(CR9R10)mNR6R7, NR7C(O)(CR9R10)rR6, NR7C(O)(CR9R10)rOR6, NR7C(xe2x95x90NC)(CR9R10)rOR6, NR7C(xe2x95x90NC)(CR9R10)rR6, NR7CO(CR9R10)rNR6R7, NR7(CR9R10)mOR6, NR7(CR9R10)rCO2R6, NR7(CR9R10)mNR6R7, NR7(CR9R10)nSO2(CR9R10)qR6, CONR7(CR9R10)nSO2(CR9R10)q, SO2NR7(CR9R10)nCO(CR9R10)qR6, SO2NR6(CR9R10)mOR6, C2-C6 alkenyl, C3-C10 cycloalkyl, C3-C10 cycloalkylmethyl, aryl, heterocyclic and alkylaryl, wherein said aryl groups may be substituted with 0-2 substituents independently selected R12;
R12 at each occurence are independently selected from H, halogen, NO2, C1-C4alkyl, C3-C10cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, oxo, C1-C4alkoxy-, OR6, O(CR9R10)rCO2R6, O(CR9R10)mNR6R7, O(CR9R10)pCN, O(CR9R10)rC(xe2x95x90O)NR6R7, C1-C4alkylcarbonyl-, CN, NH2, NHR6, NR6R7, NR7(CR9R10)rCO2R6, NR7OR6, NR7(CR9R10)mOR6, NR7CH[(CR9R10)pOR6]2, NR7C[(CR9R10)pOR6]3, NR7C(xe2x95x90O)R6, NR7(CR9R10)mOR6, NR7(CR9R10)mNR6R7, NR7(CR9R10)mSO2(CR9R10)qR6, SR7, S(O)R7, SO2R7, SO2NR6, SO3R7, CO2H, CO2R6, and CONR6R7.
Compounds of formula I, their enantiomers, diasteromers, tautomers and pharmaceutically acceptable salts, prodrugs and solvates thereof, are novel.
The present invention also provides pharmaceutical compositions comprising the compounds of formula I and methods of treating IMPDH-associated disorders using the compounds of formula I.
The compounds of the present invention offer therapeutic advantages over known prior art compounds, and are useful in treating IMPDH-associated disorders. These advantages include increased solubility (which in turn increases overall therapeutic benefit) and reduction in negative side effects.
As described above, the present invention encompasses compounds of the following formula I, and salts thereof, for use as IMPDH inhibitors: 
X is a monocyclic or bicyclic ring system optionally containing up to 4 heteroatoms selected from N, O, and S, and wherein a CH2 adjacent to any of the said N, O or S heteroatoms is optionally substituted with oxo (xe2x95x90O), and wherein X is optionally substituted by 0-3 substituents chosen from A, R1, or R2;
A is R3or R4;
R3 is a 5- or 6-membered heterocyclic ring system containing up to 4 heteroatoms selected from N, O, and S, said heterocyclic ring system being optionally substituted with 0-3 R5, wherein when R5 is hydroxy, the heterocycle may undergo tautomerization to an oxo species, or exist as an equilibrium mixture of both tautomers;
R4 is selected from H, F, Cl, Br, I, NO2, CF3, C0-C4 alkylCN, C1-C4alkoxy-, C0-C4 alkylhydroxy, C1-C4 alkyl-, C1-C4 alkylcarbonyl-, C0-C4 alkylOCOR6, C0-C4 alkylOC(xe2x95x90O)OR6, C0-C4 alkylOC(xe2x95x90O)NR6R7, NH2, NHR6, C0-C4 alkylNR6R7, C0-C4 alkylNR7C(xe2x95x90O)OR6, C0-C4 alkylNR6SO2NR6R7, C0-C4 alkylNR7SO2R6, C0-C4 alkylSR6, C0-C4 alkylS(O)R6, C0-C4 alkylSO2R6, SO3R7, C0-C4 alkylSO2NR6R7, C0-C4alkylSO2NR7CO(CR9R10)qR6, C0-C4 alkylCO2H, C0-C4 alkylCO2R6, C0-C4 alkylCONR6R7, and C0-C4CONR7SO2(CR9R10)qR6;
R5 is selected from H, halogen, NO2, C1-C4alkyl, C3-C10cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, C1-C4alkoxy, C1-C4alkylcarbonyl, CN, NH2, NHR6, NR6R7, SR7, S(O)R7, SO2R7, SO3R7, SO2NR6, CO2H, CO2R6, and CONR6R7;
R is H or C1-C4alkyl;
R1 and R2 are each independently selected from H, halogen, NO2, C1-C4alkyl, C3-C10 cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, C1-C4alkoxy-, OR6, O(CR9R10)rCO2R6, O(CR9R10)mNR6R7, O(CR9R10)pCN, O(CR9R10)rC(xe2x95x90O)NR6R7, C1-C4alkylcarbonyl-, CN, NH2, NHR6, NR6R7, NR7(CR9R10)rCO2R6, NR7OR6, NR7(CR9R10)mOR6, NR7CH[(CR9R10)pOR6]2, NR7C[(CR9R10)pOR6]3, NR7C(xe2x95x90O)R6, NR7(CR9R10)mOR6, NR7(CR9R10)mNR6R7, NR7(CR9R10)mSO2(CR9R10)qR6, SR7, S(O)R7, SO2R7, SO2NR6, SO3R7, CO2H, CO2R6, and CONR6R7; or, alternatively, R1 and R2, when on adjacent carbon atoms, maybe taken together to be methylenedioxy or ethylenedioxy;
R6, R7 and R8 are each independently selected from H, C1-C6alkyl, C3-C10 cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, C1-C6 alkylcarbonyl, C3-C7 cycloalkyl(C0-C5 alkyl)carbonyl, C1-C6 alkoxycarbonyl, aryl(C0-C5 alkyl)carbonyl, aryl(C1-C5 alkoxy)carbonyl heterocyclic(C0-C5 alkyl)carbonyl, heterocyclic(C1-C5 alkoxy)carbonyl, C1-C6alkylsulfonyl, aryllsulfonyl, heteroarylsulfonyl, C0-C4alkylaryl, C0-C4alkylheterocyclic, wherein said cycloalkyl, aryl or heterocyclic groups are substituted with 0-2 substituents independently selected from the group consisting of C1-C4alkyl, hydroxy, C1-C4 alkoxy, F, Cl, Br, haloalkyl, NO2 and CN;
or, alternatively, R6 and R7, or R6 and R8, or R7 and R8, when both substituents are on the same nitrogen atom [as in (xe2x80x94NR6R7) or (xe2x80x94NR7R8)], can be taken together with the nitrogen atom to which they are attached to form a heterocycle selected from 1-aziridinyl, 1-azetidinyl, 1-piperidinyl, 1-morpholinyl, 1-pyrrolidinyl, thiamorpholinyl, thiazolidinyl, 1-piperazinyl, 1-imidazolyl, 3-azabicyclo[3,2,2]nonan-3-yl, and 1-tetrazolyl, the said heterocycle being optionally substituted with 0-3 groups selected from oxo, C0-C4alkylOH, C0-C4alkylOC1-C4alkylCONH2, C0-C4alkylCO2alkylC0-C4, C1-C6 alkyl, C1-C4 alkoxy, C3-C7 cycloalkyl, xe2x80x94C0-C6 alkylcarbonyl, C3-C7 cycloalkylcarbonyl, C1-C6 alkoxycarbonyl, C3-C7 cycloalkoxycarbonyl, xe2x80x94NHCOalkyl, aryl, heteroaryl, aryl alkoxycarbonyl, heteroaryl alkoxycarbonyl, C1-C6 alkylsulfonyl, arylsulfonyl and heteroarylsulfonyl;
B is a monocyclic or bicyclic ring system optionally containing up to 4 heteroatoms selected from N, O, and S, and wherein a CH2 adjacent to any of the said N, O or S heteroatoms is optionally substituted with oxo (xe2x95x90O), and wherein B is optionally substituted by one to four R11 groups;
D is a monocyclic or bicyclic ring system optionally containing up to 4 heteroatoms selected from N, O, and S, and wherein a CH2 adjacent to any of the said N, O or S heteroatoms is optionally substituted with oxo (xe2x95x90O), and wherein D is optionally substituted by one to four (CR9R10)nE groups;
n is an integer having a value from 0-4;
m is an integer having a value from 2-6;
p is an integer having a value from 1-3;
q is an integer having a value from 0-3;
r is an integer having a value from 0-6;
R9 is H or C1-C4alkyl;
R10 is selected from H or C1-C4 alkyl, C1-C4 alkylhydroxy, C1-C4alkylaryl or C1-C4alkylheteroaryl, wherein said aryl or heteroaryl group may be substituted with 0-3 groups independently selected from H, halogen, NO2, C1-C4alkyl, C3-C10 cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, C1-C4alkoxy, C1-C4alkylcarbonyl, CN, NH2, NR6R7, SR6, S(O)R6, SO2R6, SO3R6, SO2NR6, CO2H, CO2R6, and CONR6R7;
R11 at each occurence are independently selected from H, halogen, NO2, C1-C4alkyl, C3-C10 cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, C1-C4alkoxy-, OR6, O(CR9R10)rCO2R6, O(CR9R10)mNR6R7, O(CR9R10)pCN, O(CR9R10)rC(xe2x95x90O)NR6R7, C1-C4alkylcarbonyl-, CN, NH2, NHR6, NR6R7, NR7(CR9R10)rCO2R6, NR7OR6, NR7(CR9R10)mOR6, NR7CH[(CR9R10)pOR6]2, NR7C[(CR9R10)pOR6]3, NR7C(xe2x95x90O)R6, NR7(CR9R10)mOR6, NR7(CR9R10)mNR6R7, NR7(CR9R10)mSO2(CR9R10)qR6, SR7, S(O)R7, SO2R7, SO2NR6, SO3R7, CO2H, CO2R6, and CONR6R7;
E is selected from H, halogen, NO2, C1-C4alkyl, C3-C10 cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, haloalkyl, haloalkoxy, OH, OR6, CN, CHO, CO2R6, CONR6R7, OCOR6, OC(xe2x95x90O)OR6, OC(xe2x95x90O)NR6R7, OCH2CO2R6, C(xe2x95x90O)R6, NH2, NHR6, NR6R7, NR7C(xe2x95x90O)R6, NR7C(xe2x95x90O)OR6, NR7C(xe2x95x90O)C(xe2x95x90O)OR6, NR7C(xe2x95x90O)C(xe2x95x90O)NR6R7, NR7C(xe2x95x90O)C(xe2x95x90O)(C1-C6alkyl), NR7C(xe2x95x90NCN)OR6, NR7C(xe2x95x90O)NR6R7, NR7C(xe2x95x90NCN)NR6R7, NR7C(xe2x80x2NR6)NR7R8, NR6SO2NR6R7, NR7SO2R6, SR6, S(xe2x95x90O)R6, SO2R6, SO3R7, SO2NR6R7, NHOH, NHOR6, NR6NR7NR8, N(COR6)OH, N(CO2R6)OH, CO2R6, CONR6R7, CONR7(CR9R10)rr6, CO(CR9R10)p O(CHR9)qCO2R6, CO(CR9CR10)rOR6, CO(CR9R10)pO(CR9R10)qR6, CO(CR9CR10)rNR6R7,OC(O)O(CR9R10)mNR6R7, O(CO)N(CR9R10)rR6, O(CR9R10)mNR6R7, NR7C(O)(CR9R10)rR6, NR7C(O)(CR9R10)rOR6, NR7C(xe2x95x90NC)(CR9R10)rR6, NR7CO(CR9R10)r NR6R7, NR7(CR9R10)mOR6, NR7(CR9R10)rCO2R6, NR7(CR9R10)mNR6R7, NR7(CR9R10)nSO2(CR9R10)qR6, CONR7(CR9R10)nSO2(CR9R10)qR6, SO2NR7(CR9R10)nCO(CR9R10)qR6, SO2NR6(CR9R10)mOR6, C2-C6 alkenyl, C3-C10 cycloalkyl, C3-C10 cycloalkylmethyl, aryl, heterocyclic and alkylaryl, wherein said aryl groups may be substituted with 0-2 substituents independently selected R12.
R12 at each occurence are independently selected from H, halogen, NO2, C1-C4alkyl, C3-C10 cycloalkyl, C2-C6alkenyl, C2-C6alkynyl, haloalkyl, haloalkoxy, OH, oxo, C1-C4alkoxy-, OR6, O(CR9R10)rCO2R6, O(CR9R10)m NR6R7, O(CR9R10)pCN, O(CR9R10)rC(xe2x95x90O)NR6R7, C1-C4alkylcarbonyl-, CN, NH2, NHR6, NR6R7, NR7(CR9R10)rCO2R6, NR7OR6, NR7(CR9R10)mOR6, NR7CH[(CR9R10)pOR6]2, NR7C[(CR9R10)pOR6]3, NR7C(xe2x95x90O)R6, NR7(CR9R10)mNR7(CR9R10)mNR6R7, NR7(CR9R10)mSO2(CR9R10)qR6, SR7, S(O)R7, SO2R7, SO2NR6, SO3R7, CO2H, CO2R6, and CONR6R7;
Preferred compounds of the present invention are compounds of the formula I, and salts thereof, wherein:
X is a monocyclic or bicyclic ring system optionally containing up to 4 heteroatoms selected from N, O, and S, and wherein a CH2 adjacent to any of the said N, O or S heteroatoms is optionally substituted with oxo (xe2x95x90O), and wherein X is optionally substituted by 0-3 substituents chosen from A, R1, or R2;
R is H;
R3 is a 5- or 6-membered heterocyclic ring system containing up to 4 heteroatoms selected from N, O, and S, said heterocyclic ring system being optionally substituted with R5, wherein when R5 is hydroxy, the heterocycle may undergo tautomerization to an oxo species, or exist as an equilibrium mixture of both tautomers;
B is a monocyclic or bicyclic ring system optionally containing up to 4 heteroatoms selected from N, O, and S, and wherein a CH2 adjacent to any of the said N, O or S heteroatoms is optionally substituted with oxo (xe2x95x90O), and wherein B is optionally substituted by one to two R11 groups;
and all other constituents are as previously described.
Particularly preferred compounds of the invention are compounds of the formula I, or salts thereof, represented by one of the following formulas II through X: 
wherein:
R3 is a 5- or 6-membered heterocyclic ring system containing up to 4 heteroatoms selected from N, O, and S, said heterocyclic ring system being optionally substituted with R5, wherein when R5 is hydroxy, the heterocycle may undergo tautomerization to an oxo species, or exist as an equilibrium mixture of both tautomers;
and all other constituents are as previously described.
In the description above and elsewhere in the specification, including the claims, each occurrence of a particular constituent is independent of each other occurrence of that same constituent.
All documents cited herein are incorporated herein by reference in their entirety.
Listed below are definitions of various terms used in the specification and claims to describe the present invention.
The term xe2x80x9calkylxe2x80x9d refers to straight or branched chain alkyl.
The term xe2x80x9cCinteger-Cintegerxe2x80x9d refers to a variable number of carbon atoms in a group depending on the integer values, as in C0-C4alkyl, which is meant to indicate a straight or branched alkyl group containing 0-4 carbon atoms. A group with 0 (zero) carbon atoms indicates that the carbon atom is absent i.e. there is a direct bond connecting adjacent terms. For example the term xe2x80x9cC0-C4 alkylhydroxyxe2x80x9d in the case xe2x80x9cC0xe2x80x9d is meant to indicate the group hydroxy.
In the case where a subscript is the integer 0 the group to which the subscript refers to indicates that the group may be absent, i.e. there is a direct bond between the groups. For example in the definition of D which may be substituted with the group xe2x80x9c(CR9R10)nE xe2x80x9cthe subscript may be 0 (zero). This is meant to indicate that D is directly connected to E by a bond i.e. Dxe2x80x94E.
The term xe2x80x9chalogenxe2x80x9d or xe2x80x9chaloxe2x80x9d refers to fluorine, chlorine, bromine or iodine.
The term xe2x80x9carylxe2x80x9d refers to monocyclic or bicyclic aromatic hydrocarbons having 6 to 12 carbon atoms in the ring portion, such as phenyl, naphthyl, biphenyl and diphenyl groups which may be optionally substituted.
The term xe2x80x9calkenylxe2x80x9d refers to straight or branched chain alkenyl groups.
The term xe2x80x9calkynylxe2x80x9d refers to straight or branched chain alkynyl.
The term xe2x80x9ccycloalkylxe2x80x9d refers to an optionally substituted, saturated cyclic hydrocarbon ring system.
The term xe2x80x9cmonocyclicxe2x80x9d or bicyclicxe2x80x9d refers to either a xe2x80x9ccarbocyclicxe2x80x9d or a xe2x80x9cheterocyclicxe2x80x9d ring system.
The term xe2x80x9ccarbocyclicxe2x80x9d refer to an optionally substituted, fully saturated or unsaturated, aromatic or nonaromatic cyclic group, which is a 3 to 7 membered monocyclic, or a 7 to 11 membered bicyclic, and all the atoms in the ring are carbon atoms. Exemplary groups include phenyl, naphthyl, anthracenyl, cyclohexyl, cyclohexenyl and the like.
The terms xe2x80x9cheterocyclexe2x80x9d and xe2x80x9cheterocyclicxe2x80x9d refer to an optionally substituted, fully saturated or unsaturated, aromatic or nonaromatic cyclic group, which is a 3 to 7 membered monocyclic, or a 7 to 11 membered bicyclic, which have at least one heteroatom and at least one carbon atom in the ring. Each heterocyclic ring may contain 1, 2, 3, or 4 heteroatoms selected from nitrogen, oxygen and sulfur, where the nitrogen and sulfur heteroatoms may also optionally be oxidized and the nitrogen heteroatoms may also optionally be quaternized. The heterocyclic group may be attached via a nitrogen or carbon atom.
Exemplary monocyclic heterocyclic groups include pyrrolidinyl, pyrrolyl, indolyl, pyrazolyl, oxetanyl, pyrazolinyl, imidazolyl, imidazolinyl, imidazolidinyl, oxazolyl, oxazolidinyl, isoxazolinyl, isoxazolyl, thiazolyl, thiadiazolyl, thiazolidinyl, isothiazolyl, isothiazolidinyl, furyl, tetrahydrofuryl, thienyl, oxadiazolyl, piperidinyl, piperazinyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, 2-oxazepinyl, azepinyl, 4-piperidonyl, pyridyl, N-oxo-pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, tetrahydrothiopyranyl, tetrahydropyranyl, morpholinyl, thiamorpholinyl, thiamorpholinyl sulfoxide, tetrahydrothiopyranylsulfone, thiamorpholinyl sulfone, 1,3-dioxolane and tetrahydro-1,1-dioxothienyl, dioxanyl, isothiazolidinyl, thietanyl, thiiranyl, triazinyl, and triazolyl, and the like.
Exemplary bicyclic heterocyclic groups include benzothiazolyl, benzoxazolyl, benzothienyl, quinuclidinyl, quinolinyl, quinolinyl-N-oxide, tetrahydroisoquinolinyl, isoquinolinyl, benzimidazolyl, benzopyranyl, indolizinyl, benzofuryl, chromonyl, coumarinyl, cinnolinyl, quinoxalinyl, indazolyl, pyrrolopyridyl, furopyridinyl (such as furo[2,3-c]pyridinyl, furo[3,1-b]pyridinyl] or furo[2,3-b]pyridinyl), dihydroisoindolyl, dihydroquinazolinyl (such as 3,4-dihydro-4oxo-quinazolinyl), benzisothiazolyl, benzisoxazolyl, benzodiazinyl, benzofurazanyl, benzothiopyranyl, benzotriazolyl, benzpyrazolyl, dihydrobenzofuryl, dihydrobenzothienyl, dihydrobenzothiopyranyl, dihydrobenzothiopyranyl sulfone, dihydrobenzopyranyl, indolinyl, isochromanyl, isoindolinyl, naphthyridinyl, phthalazinyl, piperonyl, purinyl, pyridopyridyl, quinazolinyl, tetrahydroquinolinyl, thienofuryl, thienopyridyl, thienothienyl, and the like.
xe2x80x9cIMPDH-associated disordersxe2x80x9d refers to any disorder or disease state in which inhibition of the enzyme IMPDH (inosine monophosphate dehydrogenase, EC1.1.1.205, of which there are presently two known isozymes referred to as IMPDH type 1 and IMPDH type 2) would modulate the activity of cells (such as lymphocytes or other cells) and thereby ameliorate or reduce the symptoms or modify the underlying cause(s) of that disorder or disease. There may or may not be present in the disorder or disease an abnormality associated directly with the IMPDH enzyme. Examples of IMPDH-associated disorders include transplant rejection and autoimmune disorders, such as rheumatoid arthritis, multiple sclerosis, juvenile diabetes, asthma, and inflammatory bowel disease, as well as inflammatory disorders, cancer and tumor disorders, T-cell mediated hypersensitivity diseases, ischemic or reperfusion injury, viral replication diseases, proliferative disorders and vascular diseases.
As used herein the term xe2x80x9ctreatingxe2x80x9d includes prophylactic and therapeutic uses, and refers to the alleviation of symptoms of a particular disorder in a patient, the improvement of an ascertainable measurement associated with a particular disorder, or the prevention of a particular immune response (such as transplant rejection). The term xe2x80x9cpatientxe2x80x9d refers to a mammal, preferably a human.
The compounds of this invention may contain one or more asymmetric carbon atoms and thus may occur as racemates and racemic mixtures, single enantiomers, diastereomeric mixtures and individual diastereomers. All such isomers of the compounds disclosed herein are expressly included within the scope of the present invention. Each stereogenic carbon may be of the R or S configuration.
Combinations of substituents and variables thereof that result in stable compounds are also contemplated within the present invention. The term xe2x80x9cstablexe2x80x9d as used herein refers to compounds which possess stability sufficient to allow manufacture and which maintain their integrity for a sufficient period of time to be useful as a therapeutic or diagnostic agent.
As used herein, the compounds of this invention are defined to include pharmaceutically acceptable derivatives and prodrugs thereof. A xe2x80x9cpharmaceutically acceptable derivative or prodrugxe2x80x9d includes any pharmaceutically acceptable salt, ester, salt of an ester, or other derivative of a compound of the present invention which, upon administration to a subject, is capable of providing (directly or indirectly) a compound of the invention. Particularly favored derivatives and prodrugs are those that increase the bioavailability of the compounds of the present invention when such compound is administered to a subject (e.g., by allowing an orally administered compound to be more readily absorbed into the blood) or which enhance delivery of the parent compound to a biological compartment (e.g., the brain or lymphatic system) relative to the parent species. Preferred prodrugs include derivatives where a group that enhances aqueous solubility or active transport through the gut membrane is appended to a compound of the present invention.
Pharmaceutically acceptable salts of the compounds disclosed herein include those derived from pharmaceutically acceptable inorganic and organic acids and bases known to those skilled in the art. Examples of suitable acid salts include, but are not limited to, the following: acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptanoate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oxalate, palmoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, salicylate, succinate, sulfate, tartrate, thiocyanate, trifluoroacetic, tosylate and undecanoate. Other acids, for example oxalic, while not in themselves pharmaceutically acceptable, may be employed in the preparation of salts useful as intermediates in obtaining the compounds of the present invention and their pharmaceutically acceptable acid additional salts.
Salts derived from appropriate bases include, but are not limited to, the following: alkali metal (e.g., sodium), alkaline earth metal (e.g., magnesium), ammonium and Nxe2x80x94(C1-4 alkyl)4+ salts. The present invention also envisions the quaternization of any basic nitrogen-containing groups of the compounds disclosed herein. Water- or oil-soluble or dispersible products may be obtained by such quaternization.
The compounds of the present invention may be synthesized using conventional techniques known in the art. Advantageously, these compounds are conveniently synthesized from readily available starting materials. Following are general synthetic schemes for manufacturing compounds of the present invention. These schemes are illustrative and are not meant to limit the possible techniques one skilled in the art may use to manufacture compounds disclosed herein. Different methods will be evident to those skilled in the art. Additionally, the various steps in the synthesis may be performed in an alternate sequence or order to give the desired compound(s). All documents cited herein are incorporated herein by reference in their entirety.
The group xe2x80x9cDxe2x80x9d in the synthetic schemes below is intended to designate the group in the claims defined by the letter xe2x80x9cDxe2x80x9d, and in these schemes is not intended to imply or designate the element deuterium which is often symbolized in the chemical literature by the lower case letter xe2x80x9cdxe2x80x9d, or in some chemical literature as the upper case letter xe2x80x9cDxe2x80x9d.
Compounds of the present invention can be made by many methods, which will be known to one skilled in the art of organic chemistry. In general, the time taken to complete a reaction procedure will be judged by the person performing the procedure, preferably with the aid of information obtained by monitoring the reaction by methods such as HPLC or TLC. A reaction does not have to go to completion to be useful to this invention. The preparation of heterocycles useful to this invention are described in the series of books: xe2x80x9cComprehensive Heterocyclic Chemistry. The Structure, Reactions, Synthesis and Uses, of Heterocyclic Compoundsxe2x80x9d Katritzky, A. R., Rees, C. W. Ed""s Pergamon Press New York, First edition 1984, and xe2x80x9cComprehensive Heterocyclic Chemistry II. A Review of the Literature 1982-1995. The Structure, Reactions, Synthesis and Uses, of Heterocyclic Compoundsxe2x80x9d Katritzky, A. R., Rees, C. W. and Scriven, E., F. Ed""s Pergamon Press New York, 1996. Examples of methods useful for the production of compounds of this invention are illustrated in schemes 1-24:
Appropriately substituted 1,2,4-aminotriazoles of type (II), which are useful to this invention, can be made by several means, for example as shown in scheme 1, reaction of an appropriately substituted amine with a reagent such as 1,1xe2x80x2-thiocarbonyldi-2(1H)-pyridone,1,1xe2x80x2-thiocarbonyldiimidazole or thiophosgene in a solvent such as methylene chloride or dioxane yields the isothiocyanate. Treatment of the isothiocyanate with sodium salt of cyanamide yields the sodium salt of N-cyanothiourea which is cyclized to the aminotriazole (II) using an appropriately substituted hydrazine and a dehydrating agent such as 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC), or DCC. 
1,2,4-Aminotriazoles of type (II) which are useful to this invention, may also be prepared as outlined in Scheme 2. An appropriately substituted amine can react with diphenyl cyanocarbonimidate to yield the N-cyano-O-phenylisourea. Cyclization of N-cyano-O-phenylisourea to the appropriately substituted triazole (II) is achieved using a appropriately substituted hydrazine and a solvent such as acetonitrile. 
Appropriately substituted 1,2,4-triazoles of type (III), which are useful to this invention, can be prepared as by several methods for example as outlined in Scheme 3. Acylisothiocyanates are useful intermediates in the production of some compounds of this invention and are either commercially available or readily prepared by reaction of an acid chloride, with either sodium or potassium isothiocyanate in an inert solvent such as dioxane. Acid chlorides are either commercially available or readily prepared by reaction of a carboyxylic acid and a reagent such a as thionyl chloride, or oxalyl chloride in the presence of a catalytic amount of N,N-dimethylformamide, in an inert solvent such as chloroform or methylene chloride. Reaction of an appropriately substituted amine with an acylisothiocyanate yields the thiourea. The thiourea is cyclized to (III) using hydrazine in a solvent such as ethyl alcohol or a solvent mixture such as THF and ethyl alcohol, at a temperature preferably between 60xc2x0 C. and the boiling point of the solvents utilized. 
Appropriately substituted 2-amino-1,3,4-oxadiazoles of type (IV), which are useful to this invention, can be prepared by several methods one of which is outlined in Scheme 4. Reaction of an appropriately substituted amine with a activating agent such as 1,1xe2x80x2carbonyldiimidazole followed by treatment with a appropriately substituted hydrazine yields the carbonylhydrazide. The carbonylhydrazide on treatment with 1,2-dibromotetrachloroethane and triphenylphosphine in the presence of a suitable base such as triethylamine and an appropriate solvent such as acetonitrile yields compound (IV). 
Appropriately substituted 3-amino-1,2,4 oxadiazoles of type (V), useful to this invention, can be prepared by several methods one of which is outlined in Scheme 5. Reaction of an appropriately substituted amine with a appropriately substituted acylisothiocyanate yields the thiourea. Reaction of the thiourea with a base such as sodium hydroxide or sodium hydride followed by a alkylating agent such as methyl iodide gives the S-methylisothiocarbamoyl intermediate which on treatment with hydroxylamine in the presence of a suitable solvent such as ethyl alcohol or butyl alcohol at a temperature preferably between 60 and 110xc2x0 C. results in cyclization to the desired 3-amino-1,2,4oxadiazole. 
Appropriately substituted 2-amino-1,3,4-thiadiazoles of type (VI), which are useful to this invention, can be prepared by several methods one of which is illustrated in Scheme 6. Reaction of an appropriately substituted isothiocyanate with a appropriately substituted hydrazide yields the thiourea which is cyclized to the desired heterocycle using a dehydrating agent such as methansulphonic acid, in an inert solvent such as toluene or xylene, at a temperature preferably between 80xc2x0 C. to 140xc2x0 C. 
Appropriately substituted 3-amino-1,2,4-thiadiazoles, useful to this invention, can be made by several methods known to one skilled in the art of organic chemistry. One method is outlined in scheme 7. Reaction of an appropriately substituted isothiocyanate and amidoxime in a solvent such as chloroform, or toluene at a temperature preferably between 60xc2x0 C. and 110xc2x0 C. results in the production of the desired heterocycle. Amidoximes useful to this invention are either commercially available, or readily prepared by many methods known to one skilled in the art of organic chemistry such as reaction of a nitrile with anhydrous hydrochloric acid in anhydrous methanol followed by reaction of resulting imidate with hydroxylamine. 
Appropriately substituted diarylamines, useful to this invention can be prepared by several methods include the one illustrated in Scheme 8. The coupling of amines with haloaryl compounds has been described in the chemical literature for example in xe2x80x9cRational Development of Practical Catalysts for Aromatic carbon-Nitrogen Bond Formationxe2x80x9d in Accounts of Chemical Research (1998) 31, 805-818. Reaction of an appropriately substituted aniline with an appropriately substituted haloaryl compound in the presence of a catalyst such as tris(dibenzylideneacetone)dipalladium, a ligand such as BINAP, a base such as sodium tert-butoxide and a solvent such as toluene, or dioxane, preferably at a temperature between 80-110xc2x0 C. results in production of the desired diarylamines.
Bromo or Iodo aryl intermediates such as 3-bromobiphenyl, are either commercially available or readily prepared by methods known to one skilled in the art of organic chemistry, by a variety of methods such as bromination of the aryl ring with Br2, in the presence of iron, and other methods described in chapter 11 of xe2x80x9cAdvanced Organic Chemistryxe2x80x9c3rd edition, Part B, Carey, F. A., and Sundberg, R. J., Plenum Press New York, 1990. 
3-Aminoisoxazoles which are useful to this invention, can be prepared by several methods including the method outlined in scheme 9. Reaction of an appropriately substituted isocyanate with a ketone in the presence of a base such as sodium hydride and an alkylating agent such as methyl iodide gives a thiomethyl intermediate which on treatment with hydroxylamine in a solvent such as ethyl or butyl alcohol at a temperature preferably between 60-120xc2x0 C. yields the appropriately substituted 3-aminoisoxazoles. Ketones useful to this invention are either commercially available or readily prepared by several methods such as Friedle-Crafts acylation as described in in chapter 11 of xe2x80x9cAdvanced Organic Chemistry xe2x80x9c3rd edition, Part B, Carey, F. A., and Sundberg, R. J., Plenum Press New York, 1990, hydrolysis of an enol ether, or oxidation of an alcohol, as outlined in xe2x80x9cComprehensive Organic Transformations, A Guide to Functional Group Preparationsxe2x80x9d Larock, R. C., VCH Publishers, New York, 1989. 
5-Aminothiazoles which are useful to this invention can be prepared by several methods including the method outlined in scheme 10. Reaction of an amine with a amino acid in the presence of an activating agent such as 1,1xe2x80x2-carbonyldiimidazole in a suitable solvent such as tetrahydrofuran yields an amide. Amides may be readily prepared by a number of methods including reaction of an amine with an acid chloride, or coupling of a carboxylic acid and the amine in the presence of a variety of coupling agents such as EDC, DCI, in the presence of an amine base. The coupling reaction may be enhanced by the addition of 1-hydroxybenzotriazole or similar additives. Reaction of the amide with Lawesson""s reagent in the presence of a base such as pyridine at a temperature preferably between 80-120xc2x0 C. yields the appropriately substituted 5-aminothiazoles. 
2-aminothiazoles useful to this invention can be prepared by several methods including the method outlined in scheme 11. Reaction of an isothiocyanate with ammonia in a solvent such as dioxane yields the thiourea. Aryl bromides useful to this invention, are either commercially available or readily prepared by reaction of a carboxylic acid with thionylbromide. Treatment of the thiourea with an acylbromide, in the presence of a solvent such as ethyl alcohol or dioxane, at a temperature preferably between 60xc2x0 C. and 110xc2x0 C., yields the appropriately substituted 2-amino-1,3-thiazoles. 
2-Amino 1,3-oxazolines useful to this invention may be prepared by several methods, an example of which is outlined in scheme 12. An appropriately substituted isothiocyanate is reacted with an aminoalcohol in a suitable solvent such as dioxane to yield the thiourea. Treatment of the thiourea with 2-chloro-3-ethylbenzoxazolium tetrafluoroborate in the presence of a base such as triethylamine in a solvent such as acetonitrile yields the appropriately substituted 2-amino-1,3-oxazolines.
Aminoalcohols useful to this invention are either commercially available or readily prepared by several methods. One convenient method is reduction of azidoketones of the type described in schemes 15a-15d, either by catalytic hydrogenation in the presence of palladium on carbon in a solvent such as ethanol or ethyl acetate, or by a hydride reagent such as lithium aluminum hydride in a solvent such as dioxane or tetrahydrofuran. 
2-Aminooxazoles which are useful to this invention, can be prepared by several methods, including the method outlined in Scheme 13. Reaction of a isothiocyanate with a xcex2-ketoamine in the presence of a base such as triethylamine and a solvent such as dioxane yields the thiourea. Reaction of the thiourea in the presence of a dehydrating agent such as dicyclohexylcarbodiimide or EDC, in a solvent such as dioxane or toluene, at a temperature preferably between 60xc2x0 C. and 110xc2x0 C., to yield the appropriately substituted 2-aminooxazoles. xcex2-ketoamines useful to this invention are either commercially available or readily prepared by several methods. One method is reduction of azidoketones of the type described in schemes 15a-15d, by phosphine reagents such as triphenylphosphine in a solvent such as dioxane, followed by the addition of water or dilute ammonium hydroxide. 
A second method of preparing 2-aminooxazoles is outlined in Scheme 14. Reaction of an appropriately substituted isothiocyanate with an acylazide of the type described in schemes 15a-15d in the presence of a phosphine such as triphenylphosphine in a solvent such as dichloromethane or dioxane at a temperature from room temperature to 100xc2x0 C.
Caution: Appropriate safety methods, which are known to those experienced in conducting azide reactions, such as use of a blast shield, blast wall, or similar containment device, particularly when the reaction involves heating the organic azide, as well as the use of appropriate personal protection to avoid exposure to azides which may be toxic must be exercised during the preparation and use of organic azides. 
Azides useful to this invention may be prepared using one of the sequences outlined in scheme 15a-15d.
Scheme 15a outlines the treatment of the xcex1-bromoketone with sodium azide in a solvent such as acetone, generally at room temperature, to yield the desired xcex1-azidoketones useful as intermediates in this invention. xcex1-Bromoketones are either commercially available or readily prepared by reaction of a ketone with a brominating agent such as bromine in acetic acid or pyridinium bromide perbromide and 30% hydrobromic acid. 
Scheme 15b outlines the treatment of the xcex1-bromoketone with sodium azide in acetone gives the xcex1-azidoketone. In this case, the xcex1-bromoketone is prepared by reaction of a carboxylic acid with iso-butylchloroformate and N-methylmorpholine to provide the mixed anhydride, which on treatment with diazomethane gives the xcex1-diazoketone. Reaction of the xcex1-diazoketone with either HBr gas in a solvent such as ether or dioxane, or aqueous 48% HBr, provides the xcex1-bromoketone. 
Scheme 15c illustrated preparation of xcex1-bromoketone by reaction of a ketone with sulfuric acid and bromine to yield the xcex1,xcex1-dibromoketone, which on treatment with diethylphosphite and triethylamine yields the xcex1-(mono)bromoketone. Treatment of the xcex1-bromoketone with sodium azide in acetone gives the xcex1-azidoketone. 
Scheme 15d illustrated preparation of a-bromoketone by reaction of an aryl bromide with tributyl(1-ethoxyvinyl)tin and bis-(triphenylphosphine)palladium dichloride to provide an intermediate enol ether. Treatment of the enol ether with N-bromosuccinamide at a temperature from 0xc2x0 C. to room temperature yields the xcex1-bromoketone. As previously described treatment of the xcex1-bromoketone with sodium azide in acetone gives the xcex1-azidoketone. 
Triazines useful to this invention may be prepared by many methods including the methods outlined in schemes 16-18.
Reaction of a commercially available cyanuric halide such as cyanuric chloride with an aryl Grignard reagent yields the 2-aryl substituted-4,6-dichlorotriazine. Treatment of the dichlorotriazine with an aniline in a solvent such as acetone or dioxane with or without the addition of a base such as potassium carbonate, yields the intermediate 2-arylamino-6-aryl-4-chloro triazine. The remaining chloro group on the triazine may be replaced by a variety of nucleophiles such as amines in a solvent such as dioxane at a temperature preferably between 60-140xc2x0 C., or a sodium salt of a thiol in a inert solvent or a sodium alkoxides in an appropriate alcoholic or inert solvent such as dioxane or toluene, to provide triazines of type (VIII). 
An alternative method of producing compounds useful to this invention is outlined in Scheme 17. Consecutive displacement of the chloro groups in cyanuric chloride by nucleophiles can be accomplished by careful choice of reaction conditions with particular attention to the reaction temperature and order of nucleophile addition. This has been documented in the chemical literature, for example, monosubstitution of cyanuric chloride is is depicted in Cambell, J. R., and Hatton, R. E., J. Org. Chem., 26, 2786, 1961, disubstitution of a triazine is illustrated in Thurston, J. T., Dudley, J. R, Kaiser, D. W., Schaefer, F. C, et. al. J. Amer. Chem. Soc. 73, 2981, 1954, and trisubstitution is shown in Controulis, J., Banks, C. K., J. Amer. Chem. Soc. 67, 1946, 1945. In the illustrated case, reaction of cyanuric chloride with an aniline yields the 2-arylamino-4,6-dichlorotriazine, preferably at a temperature between xe2x88x9245xc2x0 C. and room temperature. Addition of a second amine to the 2-arylamino-4,6-dichlorotriazine at an extended period of time at room temperature or preferably less than 40xc2x0 C., provides the monochlortriazine intermediate. Treatment of the monochlorotriazine intermediate with an amine at a temperature preferably between 60xc2x0 C. and 140xc2x0 C. provides the trisubstituted triazine. 
Scheme 18 illustrates another alternative preparation for triazines useful to this invention. A monochlortriazine intermediate produced in a manner similar to that described in scheme 17, is coupled with an aryl(trialkyl)tin or arylboronic acid or heteroaryl(trialkyl)tin or heteroarylboronic acid, in the presence of a palladium catalyst such as tetrakis triphenylphosphine palladium (0) to provide an aryl or heteroaryl substituted triazines of type (VIII). 
Pyrimidines useful for this invention may be prepared by several methods, one of which is illustrated in scheme 19. Reaction of an appropriately substituted aniline with 1,3 bis(tert-butoxycarbonyl)-2-methyl-2-thiopseudourea and triethylamine yields the bis-tert-butoxycarbonylguanidine. Cleavage of Boc groups using an acid such as TFA or 4N HCl in dioxane to yield the guanidine salt. There are many methods of liberating the free base of a guanidine including treatment with a base, such as sodium methoxide in anhydrous methanol, followed by filtration to remove the salt, or stirring with a commercially available strongly basic resin followed by filtration, and evaporation of the solvent. The guanidine was treated with a xcex2-ketoester and heated in a suitable solvent such as ethanol or dioxane at a temperature preferably between 50xc2x0 C.-150xc2x0 C. to yield the pyrimidinone (XI). It is recognized that more than one isomeric pyrimidone may be produced during this reaction, and the desired product may require purification by chromatography or recrystalization. Pyrimidone are also useful intermediates which can be readily converted to the chloropyrimidine by treatment with phosphoryl chloride. Displacement of the chloro group of pyrimidines can be accomplished with a variety of nucleophiles in a manner similar to that described for the triazines above. 
Imidazoles useful for this invention may be prepared according to the method outlined in scheme 20. Reaction of a guanidine (which can be obtained by the method outlined in scheme 19) with an xcex1-bromoketone yields 2-aminoimidazoles of type (X) in the presence of a base such as potassium carbonate in a solvent such as N,N-dimethylformamide provides the desired aminoimidazoles. It is recognized that more than one isomeric imidazole can form during this reaction and the desired product can be obtained by a suitable chromatographic method or by recrystalization. 
Amines attached to aryl or heteroaryl ring systems are useful as intermediates in this invention. There are many methods of preparing such intermediates known to one skilled in the art of organic chemisty. Several methods of preparing amines useful to this invention are illustrated in schemes 21-25.
A general method for the synthesis of an amine by metal catalyzed cross coupling methods is illustrated in Scheme 21, and similar examples have been reported in the chemical literature. The simplest case is a Suzuki type cross coupling (Miyaura, N., Yanagi, T. Suzuki, A., Synth. Comm. 11(7):513-519 (1981); A. Suzuki et. al., J. Am. Chem. Soc. 111:513 (1989); and V. N. Kalinin, Russ. Chem. Rev. 60:173 (1991)) of an aryl boronic acid or ester (21.1) with an appropriate bromoheterocycle in the presence of a suitable catalyst such as tetrakis(triphenylphosphine) palladium. After the cross coupling has been performed the product may be deprotected. The choice of protecting group and its method of removal will be readily apparent to one skilled in the art of organic chemistry. Such considerations and methods are, for example, described by Greene, Theodora W. and Wuts, Peter G. M. in xe2x80x9cProtective Groups in Organic Synthesis.xe2x80x9d 2nd Ed. (1991) Publisher: (John Wiley and Sons, Inc., New York, N.Y. For example, if the protecting group is acetyl the product may be deprotected by treatment with aqueous potassium hydroxide at a concentration of 0.5N to 5 N at room temperature to 100xc2x0 C. for a period between 0.5 h and 24 h, to provide amine (21.4).
For example aryl boronic acid (21.5) may react with the known 5-bromothiazole(21.6) in the presence of tetrakis(triphenylphosphine) palladium (0), to provide (21.7) which may be deprotected by an appropriate method. 
Copper has been recently been shown to be an effective catalyst for cross coupling of aryl boronic acids to N-unsubstituted heterocycles as described by Chan. et al., Tetrahed. Lett. 39:2933-2936 (1998); and Lam et al., Tetrahed. Lett. 39:2941-2944 (1998). This results in compounds in which the heterocycle is attached to the aryl ring through nitrogen rather than carbon. For example aryl boronic acid (21.5) may react with oxazolone (21.8) in the presence of copper (II) acetate in the presence of an amine base such as pyridine to provide intermediate (21.9) which may be deprotected by an appropriate method.
In general aryl boronic acids and ester, of type (21.1), where X is not Br or I, may be prepared as shown in Scheme 22, from the corresponding arylbromide 22.1 by treatment with a palladium catalyst such as [1,1xe2x80x2-Bis(diphenylphosphino)-ferrocene] dichloropalladium (II) and bis(pinacolato)diboron, (22.2), as reported by Ishayama et al., J. Org. Chem., (1995) 7508-7510. Aryl boronic esters may be converted to the corresponding boronic acid by several methods including treatment with aqueous HCl. In a variation of the synthesis, the nitrogen may be masked as a nitro group and later reduced by several means including metal reductions, such as by treatment with tin chloride in HCl or by refluxing the nitro compound with zinc in the presence of CaCl2 in a solvent such as ethanol, or in certain cases the nitro group may be reduced by catalytic hydrogenation in the presence of catalysts such as palladium on carbon. The conditions for the reduction of nitro groups are detailed in several references including Hudlicky, M., xe2x80x9cReductions in Organic Chemistryxe2x80x9d, 2nd Ed., ACS Monograph 188, 1996, pp 91-101 American Chemical Society, Washington, D.C. A second variation of the synthesis allows the aryl bromide to remain through the entire synthesis and elaborated to the boronic acid at the end. This may eliminate the need for a protecting group. 
In certain cases it may be more expedient to construct the heterocyclic ring by other methods. A general method for the synthesis of 5-membered heterocycles includes the 1,3-dipolar cycloaddition reaction, which is well known to one skilled in the art of organic chemistry and is described by Padwa, Albert; Editor. in xe2x80x9c1,3-Dipolar Cycloaddition Chemistry, Vol. 2xe2x80x9d (1984) John Wiley and Sons, New York, N.Y.; and Padwa, Albert; Editor. in xe2x80x9c1,3-Dipolar Cycloaddition Chemistry, Vol. 1xe2x80x9d (1984) John Wiley and Sons, New York, N.Y. For example oxazoles may be prepared as described in scheme 23, by 1,3 dipolar cycloaddition of the corresponding aldehyde (23.1) and (p-tolylsulfonyl)methyl isocyanate (TOSMIC) (23.2). The aldehyde may be commercially available or prepared from the corresponding methyl group by oxidation with reagents such as CrO3, MnO2, and ammonium cerium (IV) nitrate by methods well known to one skilled in the art of organic chemistry and is described in Hudlicky, M., xe2x80x9cOxidations in Organic Chemistryxe2x80x9d, ACS Monograph 186 (1990), American Chemical Society, Washington, D.C. The nitro group in intermediate (23.3) is reduced to an amine (23.4) as discussed above. 
An alternative method of producing amines useful to this invention is by nucleophilic attack an electron deficient ring system as outlined in scheme 24. Halonitrobenzenes (24.1) are either commercially available or readily prepared by methods known to one skilled in the art of organic synthesis. Displacement with a variety of nucleophiles produces compounds of structure (24.2). In one example heating (24.3) with a nucleophilic heterocycle such as triazole with or without the addition of a base provides the intermediate nitro compound which may be reduced as previously described to provide amines (24.4). Alternatively simple organic nucleophiles such as cyanide can be reacted with halonitrobenzene (24.5) to provide an intermediate nitrocompound which can be reduced by many methods to amine (25.6). 
The compounds of the present invention may be modified by appending appropriate functionalities to enhance selective biological properties. Such modifications are known in the art and include those which increase biological penetration into a given biological compartment (e.g., blood, lymphatic system, central nervous system), increase oral availability, increase solubility to allow administration by injection, alter metabolism and alter rate of excretion.
The compounds of the present invention inhibit IMPDH enzyme, and are thus useful in the treatment, including prevention and therapy of disorders which are mediated or effected by cells which are sensitive to IMPDH inhibition, as described previously. The present invention thus provides methods for the treatment of IMPDH-associated disorders, comprising the step of administering to a subject in need thereof at least one compound of the formula I, preferably at least one compound represented by formulas II through X, in an amount effective therefor. Other therapeutic agents, such as those described below, may be employed with the inventive compounds in the present methods. In the methods of the present invention, such other therapeutic agent(s) may be administered prior to, simultaneously with or following the administration of the compound(s) of the present invention.
Use of the compounds of the present invention is treating exemplified by, but is not limited to, treating a range of disorders such as: treatment of transplant rejection (e.g., kidney, liver, heart, lung, pancreas (e.g., islet cells), bone marrow, cornea, small bowel, skin allografts, skin homografts (such as employed in burn treatment), heart valve xenografts, serum sickness, and graft vs. host disease, in the treatment of autoimmune diseases, such as rheumatoid arthritis, psoriatic arthritis, multiple sclerosis, juvenile diabetes, asthma, inflammatory bowel disease (such as Crohn""s disease and ulcerative colitus), pyoderma gangrenum, lupus (systemic lupus erythematosis), myasthenia gravis, psoriasis, dermatitis, dermatomyositis; eczema, seborrhoea, pulmonary inflammation, eye uveitis, hepatitis, Grave""s disease, Hashimoto""s thyroiditis, autoimmune thyroiditis, Behcet""s or Sjorgen""s syndrome (dry eyes/mouth), pernicious or immunohaemolytic anaemia, Addison""s disease (autoimmune disease of the adrenal glands), idiopathic adrenal insufficiency, autoimmune polyglandular disease (also known as autoimmune polyglandular syndrome), glomerulonephritis, scleroderma, morphea, lichen planus, viteligo (depigmentation of the skin), alopecia areata, autoimmune alopecia, autoimmune hypopituatarism, Guillain-Barre syndrome, and alveolitis; in the treatment of T-cell mediated hypersensitivity diseases, including contact hypersensitivity, delayed-type hypersensitivity, contact dermatitis (including that due to poison ivy), uticaria, skin allergies, respiratory allergies (hayfever, allergic rhinitis) and gluten-sensitive enteropathy (Celiac disease); in the treatment of inflammatory diseases such as osteoarthritis, acute pancreatitis, chronic pancreatitis, asthma, acute respiratory distress syndrome, Sezary""s syndrome and vascular diseases which have an inflammatory and or a proliferatory component such as restenosis, stenosis and artherosclerosis; in the treatment of cancer and tumor disorders, such as solid tumors, lymphomas and leukemia; in the treatment of fungal infections such as mycosis fungoides; in protection from ischemic or reperfusion injury such as ischemic or reperfusion injury that may have been incurred during organ transplantation, myocardial infarction, stroke or other causes; in the treatment of DNA or RNA viral replication diseases, such herpes simplex type 1 (HSV-1), herpes simplex type 2 (HSV-2), hepatitis (including hepatitis B and hepatitis C) cytomegalovirus, Epstein-Barr, and human immunodeficiency virus (HIV).
Additionally, IMPDH is also known to be present in bacteria and thus may regulate bacterial growth. As such, the IMPDH-inhibitor compounds of the present invention may be useful in treatment or prevention of bacterial infection, alone or in combination with other antibiotic agents.
In a particular embodiment, the compounds of the present invention are useful for the treatment of the aforementioned exemplary disorders irrespective of their etiology, for example, for the treatment of transplant rejection, rheumatoid arthritis, inflammatory bowel disease, and viral infections.
The present invention also provides pharmaceutical compositions comprising at least one of the compounds of formula I, preferably at least one of the compounds of formulas II through X, or a salt thereof, capable of treating an IMPDH-associated disorder in an amount effective therefor, alone or in combination with at least one additional therapeutic agent, and any pharmaceutically acceptable carrier, adjuvant or vehicle. xe2x80x9cAdditional therapeutic agentsxe2x80x9d encompasses, but is not limited to, an agent or agents selected from the group consisting of an immunosuppressant, an anti-cancer agent, an anti-viral agent, an anti-inflammatory agent, an anti-fungal agent, an antibiotic, or an anti-vascular hyperproliferation compound.
The term xe2x80x9cpharmaceutically acceptable carrier, adjuvant or vehiclexe2x80x9d refers to a carrier, adjuvant or vehicle that may be administered to a subject, together with a compound of the present invention, and which does not destroy the pharmacological activity thereof. Pharmaceutically acceptable carriers, adjuvants and vehicles that may be used in the pharmaceutical compositions of the present invention include, but are not limited to, the following: ion exchangers, alumina, aluminum stearate, lecithin, self-emulsifying drug delivery systems (xe2x80x9cSEDDSxe2x80x9d) such as d(-tocopherol polyethyleneglycol 1000 succinate), surfactants used in pharmaceutical dosage forms such as Tweens or other similar polymeric delivery matrices, serum proteins such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat. Cyclodextrins such as xcex1-, xcex2- and xcex3-cyclodextrin, or chemically modified derivatives such as hydroxyalkylcyclodextrins, including 2- and 3-hydroxypropyl-xcex2-cyclodextrins, or other solubilized derivatives may also be used to enhance delivery of the compounds of the present invention.
The compositions of the present invention may contain other therapeutic agents as described below, and may be formulated, for example, by employing conventional solid or liquid vehicles or diluents, as well as pharmaceutical additives of a type appropriate to the mode of desired administration (for example, excipients, binders, preservatives, stabilizers, flavors, etc.) according to techniques such as those well known in the art of pharmaceutical formulation.
The compounds of the formula I may be administered by any suitable means, for example, orally, such as in the form of tablets, capsules, granules or powders; sublingually; buccally; parenterally, such as by subcutaneous, intravenous, intramuscular, or intrasternal injection or infusion techniques (e.g., as sterile injectable aqueous or non-aqueous solutions or suspensions); nasally such as by inhalation spray; topically, such as in the form of a cream or ointment; or rectally such as in the form of suppositories; in dosage unit formulations containing non-toxic, pharmaceutically acceptable vehicles or diluents. The present compounds may, for example, be administered in a form suitable for immediate release or extended release. Immediate release or extended release may be achieved by the use of suitable pharmaceutical compositions comprising the present compounds, or, particularly in the case of extended release, by the use of devices such as subcutaneous implants or osmotic pumps. The present compounds may also be administered liposomally.
Exemplary compositions for oral administration include suspensions which may contain, for example, microcrystalline cellulose for imparting bulk, alginic acid or sodium alginate as a suspending agent, methylcellulose as a viscosity enhancer, and sweeteners or flavoring agents such as those known in the art; and immediate release tablets which may contain, for example, microcrystalline cellulose, dicalcium phosphate, starch, magnesium stearate and/or lactose and/or other excipients, binders, extenders, disintegrants, diluents and lubricants such as those known in the art. The present compounds may also be delivered through the oral cavity by sublingual and/or buccal administration. Molded tablets, compressed tablets or freeze-dried tablets are exemplary forms which may be used. Exemplary compositions include those formulating the present compound(s) with fast dissolving diluents such as mannitol, lactose, sucrose and/or cyclodextrins. Also included in such formulations may be high molecular weight excipients such as celluloses (avicel) or polyethylene glycols (PEG). Such formulations may also include an excipient to aid mucosal adhesion such as hydroxy propyl cellulose (HPC), hydroxy propyl methyl cellulose (HPMC), sodium carboxy methyl cellulose (SCMC), maleic anhydride copolymer (e.g., Gantrez), and agents to control release such as polyacrylic copolymer (e.g., Carbopol 934). Lubricants, glidants, flavors, coloring agents and stabilizers may also be added for ease of fabrication and use.
Exemplary compositions for nasal aerosol or inhalation administration include solutions in saline which may contain, for example, benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, and/or other solubilizing or dispersing agents such as those known in the art.
Exemplary compositions for parenteral administration include injectable solutions or suspensions which may contain, for example, suitable non-toxic, parenterally acceptable diluents or solvents, such as mannitol, 1,3-butanediol, water, Ringer""s solution, an isotonic sodium chloride solution, or other suitable dispersing or wetting and suspending agents, including synthetic mono- or diglycerides, and fatty acids, including oleic acid. The term xe2x80x9cparenteralxe2x80x9d as used herein includes subcutaneous, intracutaneous, intravenous, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional and intracranial injection or infusion techniques.
Exemplary compositions for rectal administration include suppositories which may contain, for example, a suitable non-irritating excipient, such as cocoa butter, synthetic glyceride esters or polyethylene glycols, which are solid at ordinary temperatures, but liquify and/or dissolve in the rectal cavity to release the drug.
Exemplary compositions for topical administration include a topical carrier such as Plastibase (mineral oil gelled with polyethylene).
The effective amount of a compound of the present invention may be determined by one of ordinary skill in the art, and includes exemplary dosage amounts for an adult human of from about 0.1 to 500 mg/kg of body weight of active compound per day, which may be administered in a single dose or in the form of individual divided doses, such as from 1 to 5 times per day. It will be understood that the specific dose level and frequency of dosage for any particular subject may be varied and will depend upon a variety of factors including the activity of the specific compound employed, the metabolic stability and length of action of that compound, the species, age, body weight, general health, sex and diet of the subject, the mode and time of administration, rate of excretion, drug combination, and severity of the particular condition. Preferred subjects for treatment include animals, most preferably mammalian species such as humans, and domestic animals such as dogs, cats and the like, subject to IMPDH-associated disorders.
The compounds of the present invention may be employed alone or in combination with each other and/or other suitable therapeutic agents useful in the treatment of IMPDH-associated disorders, such as IMPDH inhibitors other than those of the present invention, immunosuppressants, anti-cancer agents, anti-viral agents, anti-inflammatory agents, anti-fungal agents, antibiotics, or anti-vascular hyperproliferation agents.
Exemplary such other therapeutic agents include the following: cyclosporins (e.g., cyclosporin A), CTLA4-Ig, antibodies such as anti-ICAM-3, anti-IL-2 receptor (Anti-Tac), anti-CD45RB, anti-CD2, anti-CD3 (OKT-3), anti-CD4, anti-CD80, anti-CD86, monoclonal antibody OKT3, agents blocking the interaction between CD40 and CD154 (a.k.a. xe2x80x9cgp39xe2x80x9d), such as antibodies specific for CD40 and/or CD154, fusion proteins constructed from CD40 and/or CD154/gp39 (e.g., CD40Ig and CD8gp39), inhibitors, such as nuclear translocation inhibitors, of NF-kappa B function, such as deoxyspergualin (DSG), non-steroidal antiinflammatory drugs (NSAIDs) such as ibuprofen, celecoxib and rofecoxib, steroids such as prednisone or dexamethasone, gold compounds, antiviral agents such as abacavir, antiproliferative agents such as methotrexate, leflunomide, FK506 (tacrolimus, Prograf), cytotoxic drugs such as azathiprine and cyclophosphamide, TNF-xcex1 inhibitors such as tenidap, anti-TNF antibodies or soluble TNF receptor, and rapamycin (sirolimus or Rapamune) or derivatives thereof.
The above other therapeutic agents, when employed in combination with the compounds of the present invention, may be used, for example, in those amounts indicated in the Physicians"" Desk Reference (PDR) or as otherwise determined by one of ordinary skill in the art.
The compounds disclosed herein are capable of targeting and inhibiting IMPDH enzyme. Inhibition can be measured by various methods, including, for example, IMP dehydrogenase HPLC assays (measuring enzymatic production of XMP and NADH from IMP and NAD) and IMP dehydrogenase spectrophotometric assays (measuring enzymatic production of NADH from NAD). See, e.g., Montero et al., Clinica Chimica Acta 238:169-178 (1995). Additional assays known in the art can be used in ascertaining the degree of activity of a compound (xe2x80x9ctest compoundxe2x80x9d) as an IMPDH inhibitor. The inventors used the following assay to determine the degree of activity of the compounds disclosed herein as IMPDH inhibitors:
Activity of IMPDH I and IMPDH II was measured following an adaptation of the method described in WO 97/40028. The reaction mixture was prepared containing 0.1M Tris pH 8.0, 0.1 M KCl, 3 mM EDTA, 2 mM DTT, 0.4 mM IMP and 40 nM enzyme (IMPDH I or IMPDH II). The reaction was started by the addition of NAD to a final concentration of 0.4 mM. The enzymatic reaction was followed by measuring the increase in absorbance at 340 nM that results from the formation of NADH. For the analysis of potential inhibitors of the enzyme, compounds were dissolved in DMSO to a final concentration of 10 mM and added to the assay mixture such that the final concentration of DMSO was 2.5%. The assay was carried out in a 96-well plate format, with a final reaction volume of 200 xcexcl.
The compounds disclosed herein are capable of inhibiting the enzyme IMPDH at a measurable level, under the above-described assay or an assay which can determine an effect of inhibition of the enzyme IMPDH.